摘要水通道蛋白存在于动物、植物等多种生物中。在哺乳动物中, 水通道蛋白大量存在于肾脏、血细胞和眼睛等器官中, 对体液渗透、泌尿等生理过程非常重要。在植物中, 水通道蛋白直接参与根部水分吸收及整体植株的水平衡。一个水通道蛋白分子每秒钟能够允许30亿个水分子通过,它的扩散速度远远高于简单的渗透方式。由于水通道蛋白的存在, 细胞才可以快速调节自身体积和内部的渗透压。水通道蛋白、细胞间的连丝、维管系统(导管、筛管、筛胞)对蒸腾和水分利用效率决定作用的大小, 以及与水通道蛋白基因表达调控有关的信号传递等还有待进一步深人研究。本研究以拟南芥水通道蛋白家族基因的一个成员（TIP1;1）作为研究对象，通过提取拟南芥总RNA，逆转录成互补DNA后，采用PCR扩增方法克隆TIP1;1基因，连入克隆载体中，测序验证正确后，采用酶切连接的方法，再将TIP1;1亚克隆至PVX侵染性克隆载体内，测序验证正确后即为工作质粒。将质粒线性化处理，利用T7 RNA聚合酶体外将线性化质粒转录成PVX-TIP1;1重组病毒RNA，并将其接种到4周龄的本氏烟草叶片上，观察叶片及植株表型发现。利用PVX病毒载体介导的VIGS技术，观察到接种叶片及系统叶片出现了点状的坏死斑，预示着TIP1;1与叶片衰老、坏死有关。该表型为研究进一步研究植物水通道蛋白基因的作用机制提供一个重要的线索。76777

Abstract

Aquaporin proteins exist in animals, plants and other organisms。 Found in large Numbers in mammals, the water channel protein in the kidney, blood cells and organs such as eyes, is very important for filters, urology and other physiological fluids。 In plants, water channel proteins are directly involved in the water uptake of roots and the water balance of the whole plant。 A water channel protein molecule can allow 3 billion molecules of water per second to pass, its diffusion rate is much higher than the simple way of penetration。 Due to the presence of water channel proteins, cells can quickly adjust their size and internal osmotic pressure。 Water channel protein, even the wire between the cells and vascular system (ducts, screen, sieve cells) for determining transpiration and water use efficiency, the size of the related to the water channel protein gene expression regulation and the signal transmission, it remains to be further deep research。 This research in arabidopsis water channel protein gene, a member of the family (TIP1; 1) as the research object, through the extraction of total RNA in arabidopsis thaliana, reverse transcription into a complementary DNA, TIP1 cloning by PCR amplification method; 1 gene, connected to the cloning vector, sequencing verify correct, adopt the method of enzyme digestion connection, then TIP1; 1 the clone to PVX infection sex cloning vector, sequencing after verify correct plasmid for work。 The plasmid linearized using T7 RNA polymerase in vitro transcribed into PVX-TIP1 linearized plasmids; 1 recombinant virus RNA, which was inoculated into the tobacco leaf's 4 weeks of age, were found in leaf and plant phenotype。 The use of PVX virus vector mediated VIGS, observed in inoculated leaf and system blade has punctate necrosis spot indicates that TIP1; 1 with leaf senescence and necrosis。 This phenotype provides an important clue for studying the mechanism of action of plant water channel protein gene。

    关键词：水通道蛋白，病毒诱导基因沉默，马铃薯病毒X，拟南芥，本氏烟草

Key words:  Aquaporin (AQP), Virus induce gene silencing (VIGS), Potato Virus X(PVX), Arabidopsis thaliana, Nicotiana benthamiana

目录

目录 4

引言 5

1 材料与方法